The observation, over 100 years ago, that certain chemical dyes injected into the peripheral circulation were able to gain access to most organs but not the brain led to the concept of a blood-brain barrier. Research in the 1960's demonstrated that the anatomical basis of the blood-brain barrier is the specialized endothelial cells of brain capillaries. While it has been thought that the entry of drugs, toxins, and xenobiotics into the brain is simply a function of lipophilicity, electrical charge, and molecular weight, ongoing research demonstrates that the capillary endothelium composing the blood-brain barrier is not simply an anatomic entity. A number of active transport systems exist that selectively regulate both influx and efflux of compounds across brain capillary endothelial cells. The most important drug-efflux system of the blood-brain barrier identified to date is P-glycoprotein.
P-glycoprotein, the product of the mdr1 (multidrug resistance) gene, is a 170-kD membrane-spanning, cell-surface protein that functions as a drug-efflux pump. P-glycoprotein was first identified over 20 years ago in chemotherapeutic drug-resistant tumor cells, and is now known to be a major cause of multidrug resistance in human and veterinary cancer patients. In tumor cells, P-glycoprotein functions as an ATP-dependent efflux pump resulting in decreased intracellular drug accumulation and reduced cytotoxicity. Chemotherapeutic drugs that are substrates for P-glycoprotein include Vinca alkaloids (vincristine and vinblastine), doxorubicin and related compounds, taxanes, and epipodophyllotoxins. Alkylating agents, platinum compounds, and antimetabolites are not substrates for P-glycoprotein. Though these agents are structurally and functionally dissimilar, P-glycoprotein substrates share several other characteristics. They typically are complex, hydrophobic, amphipathic compounds that are natural products (i.e., derived from plants or micro-organisms) or analogs of natural products. A number of non-cytotoxic compounds have been identified as P-glycoprotein substrates, including steroid hormones, bilirubin, antiparasitic agents, selected antimicrobial agents, and others.
P-glycoprotein is expressed not only in tumor cells, but also in a variety of normal tissues, including renal tubular epithelium, canalicular surfaces of hepatocytes, adrenal cortical cells, colonic and intestinal epithelium, placenta, apical margins of bronchiolar epithelium, and brain capillary endothelial cells. Consistent with its function as a transport pump, the expression of P-glycoprotein in non-neoplastic tissues suggests a normal physiologic role for P-glycoprotein mediating the export of potentially toxic xenobiotics from the body. Although the normal function of P-glycoprotein in many of these tissues has not been elucidated, a great deal is known about its role in the blood-brain barrier.
Avermectins are a class of natural products with broad antiparasitic activity. Ivermectin, a semi-synthetic lactone in the avermectin family, is a drug that is used extensively in veterinary medicine to treat and control infections caused by nematode and arthropod parasites. It is also used in human medicine to treat onchocerciasis, lymphatic filariasis, and strongyloidiasis. Ivermectin induces a tonic paralysis in invertebrate organisms by potentiating glutamate-gated chloride channels, and/or gamma-amino butyric acid (GABA)-gated chloride channels (Tracy and Webster, In: Goodman & Gilman's The Pharmacological Basis of Therapeutics, 9th edition. Hardman et al., eds. New York: McGraw-Hill, 1996: 1009-1026, 1996) of the peripheral nervous system. In most mammals, the blood-brain barrier prevents access of ivermectin to the central nervous system, and since GABA receptors in mammals are restricted to sites within the central nervous system, mammals are generally protected from the neurologic effects of ivermectin (Fisher and Mrozik, Annu. Rev. Pharmacol. Toxicol. 32:537-553, 1992).
There are some specific subgroups of mice and dogs, however, that are exquisitely sensitive to the neurologic actions of ivermectin. Genetically engineered mdr1a knock-out [mdr1a(−/−)] mice are 50 to 100 times more sensitive to ivermectin-mediated neurotoxicity than wild-type mice (Schinkel et al., Cell. 77:491-502, 1994), and accumulate 80-90-fold higher concentrations of ivermectin in the brain than do wild-type mice. The protein product of mdr1a, called P-glycoprotein (P-gp) is a 170-kD transmembrane protein pump that is present at high concentrations in the apical membrane of brain capillary endothelial cells (Van Asperen et al., J. Pharmaceut. Sci. 86:881-884, 1997, 1997; Tsuji, Therap. Drug Monitor. 20:588-590, 1998). Substrates of P-gp include a variety of large, structurally unrelated hydrophobic compounds, including naturally occurring compounds such as ivermectin, cyclosporin, digoxin, and others. After substrates are bound by P-gp, they are actively extruded from the endothelial cell into the capillary lumen (Van Asperen et al., J. Pharmaceut. Sci. 86:881-884, 1997). Abrogation of P-gp results in failure of the blood-brain barrier. High concentrations of ivermectin accumulate in brain tissue from mdr1a (−/−) mice, and neurotoxicity ensues.
Approximately 25% of a population of the CF-1 mouse strain were much more sensitive to neurotoxicity produced by ivermectin than unaffected mice of the same strain (Umbenhauer et al., Toxicol Appl. Pharmacol. 146:88-94, 1997). Investigation into the cause of this sensitivity led to the discovery that the sensitive animals did not express P-gp in their brain endothelial cells. Furthermore, a restriction-fragment-length polymorphism in the murine mdr gene was documented that allowed prediction of sensitive animals, and an inheritance pattern following normal Mendelian genetics was observed (Umbenhauer et al., Toxicol Appl. Pharmacol. 146:88-94, 1997).
In dogs, a breed-related sensitivity to ivermectin has been reported in Collies, that may affect 30 to 50% of the population (Pulliam et al., Veter. Med. 7:33-40, 1985; Hsu et al., Comp. Contin. Educat. Veter. Pract. 11:584-589, 1989, Paul et al., Am. J. Vet. Res. 48:685-688, 1987). In one study, 1/200th of the lethal dose of ivermectin for Beagles was lethal for Collies (Pulliam et al., Veter. Med. 7:33-40, 1985). Other related canine breeds believed to be affected by ivermectin sensitivity include Border Collies, Shetland Sheepdogs, Old English Sheepdogs, and Australian Shepherds (Campbell and Benz, J. Vet. Pharmacol. Therap. 7:1-16, 1984).
Despite numerous investigations (Vaughn, et al., Vet. Res. Commun. 13:47-55, 1989; Roher et al., Vet. Res. Commun. 14:157-165, 1990; Pulliam et al., Veter. Med. 7:33-40, 1985), the mechanism for ivermectin-sensitivity in Collies is unknown.